Regularly performed endurance training results in a two-fold greater density of skeletal muscle mitochondria. Aminolevulinic acid synthase (AS), the rate limiting enzyme in heme biosynthesis, is increased two-fold in skeletal muscle after a single exhaustive bout of exercise in untrained rats and precedes any change in cytochrome c content. These results suggest an important role for AS induction in the regulation of mitochondrial cytochrome production with endurance training. The primary goals of our research will be to determine (i) the isoform of AS expressed in fast and slowtwitch skeletal muscle, (ii) the pattern of changes in AS enzyme content following daily bouts of exercise in relation to selected parameters of mitochondrial biogenesis, (iii) the mechanism of AS induction by exercise, and (iv) effects of heme on skeletal muscle AS synthesis and mRNA expression. To accomplish these goals we will produce an antibody specific for AS to enable measurement of AS enzyme content and synthesis, and obtain an oligonucleotide probe from rat AS cDNA to measure AS mRNA content. We will also use the oligonucleotide to perform primer extension of various muscle RNAs to conclusively identify the isoform of AS expressed. Following one or several bouts of daily exercise, the pattern of changes in AS enzyme and mRNA content will be determined and compared to levels of cytochrome c, mitochondrial DNA, and isocitrate dehydrogenase. Rates of AS synthesis will also be correlated to mRNA contents at selected time points during induction by exercise. The probable role of intracellular heme levels in regulating AS synthesis and mRNA expression will be tested at rest using drug modulation of heme synthesis. Finally, we will isolate the gene for rat AS to eventually determine the sequence elements involved in AS enzyme induction. The results of this work may establish one of the pathways utilized by muscle cells to effect an overall increase in mitochondrial density due to daily endurance exercise training. The mechanisms studied may also be part of a more general adaptive scheme by which anemeia, hypoxia, thyrotoxicosis, and hypertrophy lead to changes in skeletal and cardiac muscle mitochondrial biogenesis.